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RARα (Retinoic Acid Receptor Alpha) is a member of the nuclear receptor superfamily. Its structure comprises a highly conserved DNA-binding domain (DBD—containing zinc-finger motifs) and a ligand-binding domain (LBD—containing the ligand-dependent transcriptional activation region AF-2). Acting as a heterodimer (in association with RXR), it specifically recognizes Retinoic Acid Response Elements (RAREs) located within the promoters of target genes. Upon binding of the endogenous ligand, all-trans retinoic acid (ATRA), to the LBD, a conformational change is triggered; this leads to the dissociation of co-repressor complexes (e.g., N-CoR/SMRT) and the recruitment of co-activators (e.g., CBP/p300), thereby regulating key pathways such as embryonic development (including neural crest differentiation and limb formation), cellular proliferation and differentiation (e.g., myeloid hematopoietic stem cell lineage commitment), and apoptosis. 

RARα Reporter Cells are recombinant cell lines that stably express human RARα, wherein the expression of a reporter gene is regulated by specific regulatory elements. Retinoic acid serves as the natural agonist for RAR nuclear receptors; the underlying principle is illustrated in the figure below.

Figure 1. Schematic Diagram of the RARα Reporter Cell Mechanism

Figure 2. Dose Response of Retinoic acid in RARα Reporter Cell(C4).

Figure 3. Inhibition of Retinoic acid induced Reporter Activity by RARα Antagonist in RARα Reporter Cell(C4).

Cell Resuscitation
1)Rapidly thaw the frozen cells in a 37 °C water bath for approximately 60 seconds. Once thawed (which may take slightly less or more than 60 seconds), immediately transfer the cell suspension from the cryovial into a 15 mL centrifuge tube containing 10 mL of pre-warmed HEK293 Human RARα Reporter Cell complete culture medium.
2)Centrifuge cells at 1000 rpm for 5 min to remove medium, then resuspend cells in 5 mL of pre-warmed complete medium.
3)Transfer the cell suspension into a T25 culture flask and incubate at 37 °C with 5% CO₂.
4)After approximately 24–36 hours, replace the medium or passage the cells to remove non-adherent dead cells.

Subculturing procedure
1)When the cell density reaches the appropriate confluency for passaging, wash the cells with PBS, then add 1 mL trypsin to detach the cells. When more than 80% of the cells detach upon gently tapping the culture flask, add complete culture medium to terminate digestion. Gently pipette to obtain a single-cell suspension, transfer to a 15 mL centrifuge tube, and centrifuge at 1000 rpm for 5 minutes.

2)Discard supernatant after centrifugation. Resuspend cells in fresh medium to a single-cell suspension and transfer to a new culture flask for continued growth.

Cell Freezing
After trypsinization and centrifugation of cells from each T75 flask or 10 cm culture dish, discard the supernatant. Add 2 mL of cryopreservation medium (90% FBS + 10% DMSO), gently resuspend thoroughly, and aliquot into two cryovials. Immediately place the cryovials into a controlled-rate freezing container (e.g., Nalgene 5100-0001), fill with isopropanol to the indicated level, and store at −80 °C. After 24 hours, transfer the cryovials to liquid nitrogen for long-term storage.